Aquaglyceroporins form a subfamily of the aquaporin water channel family-transporting water, glycerol and urea. Human red blood cells [RBCs] contain the aquaglyceroporin AQP3;mouse RBCs contain AQP9. Plasmodia causing malaria contain aquaglyceroporins -- P. falciparum causes human malaria and expresses PfAQP;P. berghei causes mouse malaria and expresses PbAQP. Glycerol is used by the intracellular malaria parasite [merozoite] for production of glycerolipids. Import of glycerol requires transport of the solute across three membranes - the RBC plasma membrane [PM], the parasitophorous vacuolar membrane [PVM], and the plasmodia plasma membrane [PPM]. We hypothesize that aquaglyceroporins are involved in malarial infection and may provide a new pathway of potential therapeutic importance. Aim I. To understand the contribution of aquaglyceroporins to glycerol transport by RBC membranes, the copy numbers will be determined for AQP3 in human and AQP9 in mouse RBCs. We will also compare the water, glycerol, and urea permeability of aquaglyceroporin null RBCs to wildtype cells. Aim II. The biophysical functions of aquaglyceroporin PbAQP from P. berghei will be characterized. In addition the localization and expression of PbAQP will be defined during the life cycle of the organism. Aim III. Aquaglyceroporin null cells will allow us to understand the role of these channels in malarial infection. The pathogenicity of PbAQP null P. berghei will be compared to that of wildtype parasites. The role of AQP9 will be determined by comparing the parasitemia of wildtype mice to that of AQP9 null mice. The mouse model of malaria will also be evaluated in the insect stages of Plasmodium by studying the expression of PbAQP in the insect and the effect of PbAQP disruption on the proliferation of P. berghei in the mosquito and the transmission of P. berghei from the mosquito to the mouse. Aim IV. Investigation of the human malaria parasite, P. falciparum, will be crucial for understanding roles for aquaglyceroporins in human malarial infection. Human RBCs will be infected with PfAQP null or wildtype P. falciparum, and the parasitemia compared. To determine the role of the human RBC aquaglyceroporins in malaria, parasitemia of wildtype RBCs will be compared to AQP3 null RBCs. Malaria is a major cause of disease and death of children in many underdeveloped countries. The parasites causing malaria invade red blood cells and multiply, causing massive cellular destruction. The goal of this application is to define the pathway for uptake of the nutrient glycerol by red blood cells and malaria parasites, with hope that this may reveal new avenues for prevention or treatment of malaria.